The overall goals of this proposal are to define the structural basis of voltage sensor movement and the mechanism of coupling sensor movement to opening of HERG delayed rectifier K+ channels. HERG is one of several voltage-gated IC channels that mediate repolarization of the cardiac action potential. HERG channel dysfunction, caused by mutations or commonly prescribed medications, is associated with life-threatening arrhythmias. The critical role of HERG in the maintenance of normal cardiac electrical activity derives from its unusual gating properties: slow channel activation and fast inactivation. Activation and inactivation gating are voltage dependent, implying that these processes are coupled to movement of charged residues within the membrane. The structural basis of voltage sensor movement and mechanism of coupling sensor movement to HERG channel opening are not known. In this proposal, we provide the first description of HERG voltage sensor movement as measured by gating current. We hypothesize (1) the domains surrounding the S4 domain (the putative voltage sensor) influence the rate of S4 movement and account for the slow activation of HERG ionic current, (2) inactivation and activation gating are coupled to the same fundamental voltage sensing mechanism and (3) voltage sensor movement is coupled to channel opening via a direct interaction between the intracellular S4-S5 linker and the S6 domain. The aims of this proposal are to determine the structural basis of slow activation in HERG channels, to define the voltage sensor movement associated with fast HERG inactivation and to determine the mechanism whereby movement of the voltage sensor is coupled to channel opening. These aims will be accomplished using site-directed mutagenesis, chimeric constructs between HERG and the structurally related EAG channel, biochemical assays, two microelectrode voltage clamp and cut-open oocyte voltage clamp techniques. Insight into the molecular basis of voltage sensing in the HERG channel will advance our understanding of how this channel contributes to normal electrical stability in the heart and may advance our understanding of arrhythmia susceptibility.